The type 1 insulin-like growth factor receptor (IGF1R) is a potent proto-oncogene implicated in the development and progression of a large number of human malignancies through its powerful mitogenic, anti- apoptotic, and transforming activities. Our lab has determined that IGF1R gene expression is regulated at the translational level through an internal ribosome entry site (IRES) contained within the 5'-untranslated region of the mRNA. Our research has focused on determining how the IRES operates, how it is regulated by sequence-specific RNA-binding proteins, and how these molecules and mechanisms may be altered in cancer. Our results led us to hypothesize that a small molecule inhibitor of the IGF1R IRES could be developed which could be used to specifically block IGF1R protein synthesis and thereby reverse the survival / proliferative advantage associated with IGF1R overexpression in tumor cells. An IRES-targeted drug potentially represents a gene-specific translational inhibitor, with enormous potential utility, both for research and therapeutic applications. Toward this objective, we have initiated a major collaborative effort with Southern Research Institute to perform a high throughput screen of a 125,000 compound library to identify compounds capable of inhibiting translation mediated through the IGF1R IRES. The Specific Aims are: 1. Utilize a series of mechanistic and functional biological assays to characterize the IRES inhibitors emanating from the high throughput screen, analyzing the consequences for IGF1R signaling while leaving insulin receptor signaling intact, and assessing the potential of these compounds to induce favorable phenotypic changes in breast tumor cells, including decreased capacity for clonogenic proliferation and enhanced sensitivity to cytotoxic chemotherapeutic agents. 2. Test the anti-tumor and anti-metastatic efficacy of the most promising IRES inhibitors in animal models, using serial in vivo bioluminescence imaging to correlate changes in IRES activity with progression or regression of the tumor in real time. 3. Perform iterative synthetic modifications to selected lead compounds emerging from the compound progression pathway, coordinated with biological re-testing and structure-activity relationship analyses, to optimize efficacy and drug-like properties of the IRES inhibitors. 4. Investigate the manner in which the newly identified IGF1R IRES-regulatory proteins (ITAFs) collectively modulate IGF1R translational efficiency, how the RNA-binding activities of these IRES- regulatory proteins may be altered in primary human breast tumors and metastatic lesions, and how specific interactions between these IRES-regulatory proteins, the IGF1R IRES, and the 40S ribosome may be blocked by the small molecule IRES inhibitors. PUBLIC HEALTH RELEVANCE: These studies are designed to advance our understanding of how gene-specific translational regulation is accomplished in the cell, and how these regulatory mechanisms may be altered in malignant cells. The IRES- targeted compounds we seek to develop would represent the first of an essentially unprecedented new category of anti-cancer drugs, with an entirely novel mechanism of action.